Polarimetry

ABSTRACT

The present invention relates to an assembly and method for measuring the optical activity of a stimulus of interest. The assembly comprises a source of incident electromagnetic radiation ( 1 ), a lens arrangement ( 2 ), an input polariser ( 3 ), a planar waveguide structure ( 4 ) (or an optical fiber), an output polariser ( 5 ), a lens arrangement ( 6 ) for focussing the output electromagnetic radiation and a detector. The polarisation of the incident electromagnetic radiation relative to the planar waveguide structure ( 4 ) is determined by the input polariser ( 3 ). The planar waveguide structure in this embodiment comprises a silicon substrate layer ( 4   c ) and an absorbent layer ( 4   e ). The silicon oxynitride layer ( 4   c ) acts as the reference waveguide and the absorbent layer ( 4   e ) acts as the sensing waveguide. The sensing waveguide ( 4   e ) is exposed in the localised environment ( 8 ) to a smaple containing the stimulus of interest which is optically active. The adjusted attitude of the output polariser ( 5 ) is directly proportional to the optical activity the stimulus of interest.

The present invention relates to an assembly and method for measuringthe optical activity of a stimulus of interest.

Measurements of the degree of rotation of plane polarised light by asample as a function of wavelength and concentration are important ininter alia the field of biology and biological research. Suchmeasurements of optical activity have formed the basis of opticalpolarimetry utilised to assess chirality and optical isomerism for overa century and remain important as (in most other respects) opticalisomers have identical properties which in general makes them difficultto distinguish or separate. For example, such measurements enable abiologically active optical isomer to be distinguished from itscomplementary optical isomer which may be biologically inactive.

Techniques relating to optical activity have been extended to includeinter alia circular dichroism (CD) spectroscopy in which the behaviourof circularly polarised light is analysed at different wavelengths. Whensuch wavelengths are in the UV region, important data relating (forexample) to the structure and folding of proteins can be deduced. Thisis because the different tertiary structures adopted by proteinmolecules (especially alpha helical and beta pleated sheets) tend to beasymmetrical and have intrinsic optical properties which can bedifferentiated in terms of their optical activity. Whilst such data isvery useful, its analysis is subject to some ambiguity.

The present invention seeks to improve the measurement of the opticalactivity of chemical or biological stimuli by exploiting the response ofcertain optical components such as waveguide structures to which thestimuli are exposed.

Thus viewed from one aspect the present invention provides an assemblyfor measuring the optical activity of a stimulus of interest in alocalised environment, said assembly comprising:

-   -   a source of incident electromagnetic radiation;    -   an input polariser for orienting the incident electromagnetic        radiation into a first degree of polarisation;    -   an optical component capable of exhibiting a measurable response        to a change in the degree of polarisation of the incident        electromagnetic radiation into output electromagnetic radiation        having a second degree of polarisation caused by the        introduction of or changes in the stimulus of interest in the        localised environment;    -   means for measuring the measurable response; and    -   means for relating the measurable response to the optical        activity of the stimulus of interest.

The optical activity of the stimulus of interest measured in accordancewith the invention may be advantageously used to deduce its structural,conformational, biological or chemical properties (or changes therein).For example, the optical activity may be used to deduce the chirality ofthe stimulus of interest.

The assembly of the invention may advantageously be used to measure theoptical activity of a chemical stimulus contained in an analyte which isintroduced into the localised environment. For example, a gaseous,vapour or liquid phase analyte comprising chemical stimuli may beintroduced into the localised environment. Alternatively, a chemicalreaction or interaction may take place which effects changes in thenature of the chemical stimuli in situ (ie causes a change in thelocalised environment) which can be deduced from changes in the opticalactivity measured in accordance with the invention.

The assembly of the invention may advantageously be used to measure theoptical activity of a biological stimulus such as an enzyme, DNAfragment, protein, antibody or whole cell contained in an analyte whichis introduced into the localised environment. Alternatively a change inthe localised environment may be brought about by (for example)conformational changes in a biological stimulus (eg a change in thefolding of a protein) which can be deduced from changes in the opticalactivity measured in accordance with the invention. Such conformationalchanges may be caused by (for example) an increase in temperature.

The stimulus of interest in the localised environment may be close to,in contact with, immobilised or bound to the optical component in anyconvenient manner.

The input polariser may be an input plane polariser (ie for orientingthe incident electromagnetic radiation into a first degree of planepolarisation). For example, the input polariser may be a polarisingfilter or prism.

Preferably the assembly further comprises: a lens arrangement capable offocussing the incident electromagnetic radiation.

Preferably the assembly further comprises: a lens arrangement capable offocussing the output electromagnetic radiation.

Preferably the means for relating the measurable response to the opticalactivity of the stimulus of interest comprises:

-   -   an output polariser for orienting the degree of polarisation of        the output electromagnetic radiation.

Preferably the output polariser is an output plane polariser. The outputpolariser may comprise a prism.

The optical component may be an optical fibre. Typically an opticalfibre would be enclosed in cladding and the localised environment may becreated by removing a portion of the cladding to expose the fibresurface.

In a preferred embodiment, the optical component is a polarisationmaintaining optical component. An example is a polarisation maintainingoptical fibre.

In a preferred embodiment, the optical component is a waveguidestructure such as a waveguide structure of the type disclosed inWO-A-98/22807 or WO-A-01/36945. The waveguide structure preferablyincludes either (1) one or more sensing layers capable of inducing in asecondary waveguide a measurable response to a change in the degree ofpolarisation of the incident electromagnetic radiation into outputelectromagnetic radiation having a second degree of polarisation causedby the introduction of or changes in the stimulus of interest in thelocalised environment or (2) a sensing waveguide capable of exhibiting ameasurable response to a change in the degree of polarisation of theincident electromagnetic radiation into output electromagnetic radiationhaving a second degree of polarisation caused by the introduction of orchanges in the stimulus of interest in the localised environment.

The measurable response may be or relate to a change in the intensity(eg the total intensity) of the output electromagnetic radiation. Forexample, this may be measurable by coupling the output electromagneticradiation into free space and generating an interference pattern. Theinterference pattern may be measured in a conventional manner (see forexample WO-A-98/22807). For example, the interference fringes may bemeasured either using a single detector which measures changes in theoutput electromagnetic radiation intensity or a plurality of suchdetectors which monitor the changes in intensity occurring in a numberof fringes or the entire interference pattern. The one or more detectorsmay be one or more photodetectors. For the most sensitive performance,the (or each) photodetector is a photomultiplier tube capable ofcounting photons. Where more than one photodetector is used this may bearranged in an array.

Preferably the assembly of the invention wherein the optical componentis a waveguide structure is adapted so as to be usable in evanescentmode or whole waveguide mode.

Thus in a first preferred embodiment of the invention, the opticalcomponent includes one or more sensing layers capable of inducing in asecondary waveguide a measurable response to a change in the degree ofpolarisation of the incident electromagnetic radiation into outputelectromagnetic radiation having a second degree of polarisation causedby the introduction of or changes in the stimulus of interest in thelocalised environment. In this first embodiment, the assembly isadvantageously adapted to optimise the evanescent component.

Preferably the (or each) sensing layer comprises an absorbent material(eg a polymeric material such as polymethylmethacrylate, polysiloxane,poly-4-vinylpyridine) or a bioactive material (eg containing antibodies,enzymes, DNA fragments, functional proteins or whole cells). Theabsorbent material may be capable of absorbing (or capturing) a gas, aliquid or a vapour analyte containing a chemical stimulus of interest,The (or each) sensing layer may comprise a porous silicon materialoptionally biofunctionalised with antibodies, enzymes, DNA fragments,functional proteins or whole cells.

In a preferred assembly of the invention, the secondary waveguidecomprises silicon oxynitride or silicon nitride.

In a second preferred embodiment of the invention, the optical componentincludes a sensing waveguide capable of exhibiting a measurable responseto a change in the degree of polarisation of the incidentelectromagnetic radiation into output electromagnetic radiation having asecond degree of polarisation caused by the introduction of or changesin the stimulus of interest in the localised environment. In this secondembodiment, the assembly is adapted to minimise the evanescent componentand may be used advantageously in whole waveguide mode.

Preferably the sensing waveguide comprises an absorbent material (eg apolymeric material such as polymethylmethacrylate, polysiloxane,poly-4-vinylpyridine) or a bioactive material (eg containing antibodies,enzymes, DNA fragments, functional proteins or whole cells). Theabsorbent material may be capable of absorbing (or capturing) a gas, aliquid or a vapour analyte containing a chemical stimulus of interest.The sensing waveguide may comprise a porous silicon material optionallybiofunctionalised with antibodies, enzymes, DNA fragments, functionalproteins or whole cells.

Where the optical component of the assembly of the invention comprises asensing waveguide adapted for use in whole waveguide mode, an absorbentlayer in the form of an overcoating may be present for use as a membrane(for example) to separate out stimuli of interest.

To optimise the performance of the first preferred embodiment, theoptical component may further comprise an inactive secondary waveguidein which the sensing layer is incapable of inducing a measurableresponse to a change in the degree of polarisation of the incidentelectromagnetic radiation into output electromagnetic radiation having asecond degree of polarisation caused by the introduction of or changesin the stimulus of interest in the localised environment. The inactivesecondary waveguide is capable of acting as a reference layer. It ispreferred that the secondary waveguide and inactive secondary waveguidehave identical properties with the exception of the measurable responseto a change in the degree of polarisation of the incidentelectromagnetic radiation into output electromagnetic radiation having asecond degree of polarisation caused by the introduction of or changesin the stimulus of interest in the localised environment. By way ofexample, the secondary waveguide and inactive secondary waveguide aremade of silicon oxynitride.

To optimise the performance of the second preferred embodiment, theoptical component may further comprise an inactive (eg deactivated)waveguide substantially incapable of exhibiting a measurable response toa change in the degree of polarisation of the incident electromagneticradiation into output electromagnetic radiation having a second degreeof polarisation caused by the introduction of or changes in the stimulusof interest in the localised environment. The inactive waveguide iscapable of acting as a reference layer. The physical, biological andchemical properties of the sensing waveguide and inactive waveguide areas similar as possible (with the exception of the response to a changein the degree of polarisation of the incident electromagnetic radiationinto output electromagnetic radiation having a second degree ofpolarisation caused by the introduction of or changes in the stimulus ofinterest in the localised environment). Typically the inactive waveguideis made of silicon oxynitride.

Preferably each of the sensing waveguide and secondary waveguide (and ofany additional waveguides such as a reference waveguide) of the opticalcomponent is a planar waveguide (ie a waveguide which permits lightpropagation in any arbitrary direction within the plane).

Preferably the waveguide structure constitutes a multi-layered structure(eg a laminated waveguide structure). In this sense, the waveguidestructure is simple to fabricate and fault tolerant in terms ofconstruction errors. In a preferred embodiment, the plurality of layersin the multi-layered optical component are built onto a substrate (eg ofsilicon) through known processes such as vapour deposition (eg PECVD orLPCVD). Such processes are highly repeatable and lead to accuratemanufacture. Intermediate transparent layers may be added (eg silicondioxide) if desired. Typically the optical component is a multilayeredstructure of thickness in the range 0.2-10 microns. A layered structureadvantageously permits layers to be in close proximity (eg a sensingwaveguide and an inactive (reference) waveguide may be in closeproximity to one another so as to minimise the deleterious effects oftemperature and other environmental factors). Preferably, the opticalcomponent comprises a stack of transparent dielectric layers whereinlayers are placed in close proximity. Preferably each layer isfabricated to allow equal amounts of electromagnetic radiation topropagate by simultaneous excitation of the guided modes in thewaveguide structure. Particularly preferably the amount ofelectromagnetic radiation in the sensing waveguide/inactive waveguide orin the secondary waveguide/inactive secondary waveguide is equal.

The assembly may further comprise: means for intimately exposing atleast a part of the optical component (eg at least a part of the (oreach) sensing layer or the sensing waveguide) to the localisedenvironment (eg as described in WO-A-01/36945). For example, the meansfor intimately exposing at least a part of the optical component to thelocalised environment may be a part of a microstructure positionable onthe surface of and in intimate contact with the optical component. Themicrostructure may comprise means for intimately exposing at least apart of the optical component to the localised environment in the formof one or more microchannels and/or microchambers. For example, ananalyte containing chemical stimuli may be fed through microchannels orchemical reactions may take place in an analyte located in amicrochamber. An analyte containing chemical stimuli may be fed into themicrochannels by capillary action or positively fed by an urging means.The means for intimately exposing at least a part of the opticalcomponent to the localised environment may be integrated onto theoptical component.

The means for intimately exposing at least a part of the opticalcomponent to the localised environment may be adapted to induce chemicalor biological changes in a static analyte containing a chemical orbiological stimulus of interest. In this sense, the system may beconsidered to be dynamic. Chemical or biological changes (eg reactions)may be induced in any conventional manner such as by heat or radiation.

The means for intimately exposing at least a part of the (or each)sensing layer or the sensing waveguide to the localised environment maybe included in a cladding layer. For example, microchannels and/ormicrochambers may be etched into the cladding layer. The cladding layermay perform optical functions such as preventing significantdiscontinuities at the boundary of the sensing waveguide or sensinglayer(s) or chemical functions such as restricting access of certainspecies to the sensing waveguide or sensing layer(s). The cladding layermay be integrated onto the optical component.

Incident electromagnetic radiation generated by the source of incidentelectromagnetic radiation may be propagated into the optical componentin a number of ways. For example, incident electromagnetic radiation maybe input via an end face of the optical component (this is sometimesdescribed as “an end firing procedure”). Preferably, the source ofincident electromagnetic radiation provides incident electromagneticradiation having a wavelength falling within the visible or UV range.Propagating means may be employed for substantially simultaneouslypropagating incident electromagnetic radiation into a plurality ofwaveguides. For example, one or more coupling gratings or mirrors may beused. A tapered end coupler rather than a coupling grating or mirror maybe used to propagate light into the lowermost waveguide.

As a consequence of the introduction of a biological and/or chemicalstimulus in the localised environment (ie a change in the refractiveindex of material in the localised environment), changes in thedielectric properties (eg the effective refractive index) of the opticalcomponent (eg sensing waveguide or sensing layer) occur. This causes achange in the transmission of incident electromagnetic radiation downthe optical component (eg sensing waveguide (or waveguides) in wholewaveguide mode or the secondary waveguide in evanescent field mode)which manifests itself as movement of the fringes in the interferencepattern.

In an embodiment of the assembly, the source of incident electromagneticradiation and one or more detectors are integrated into the assembly.

Viewed from a yet further aspect the present invention provides a methodfor measuring the optical activity of a stimulus of interest in alocalised environment, said method comprising the steps of:

-   -   (A) providing an assembly as hereinbefore defined;    -   (B) irradiating the optical component with incident        electromagnetic radiation of a first polarisation;    -   (C) measuring a first characteristic of the output        electromagnetic radiation;    -   (D) introducing to the localised environment a stimulus of        interest;    -   (E) measuring a second characteristic of the output        electromagnetic radiation; and    -   (F) relating the change from the first characteristic of the        output electromagnetic radiation to the second characteristic of        the output electromagnetic radiation to the optical activity of        the stimulus of interest.

In a preferred embodiment, the first and second characteristic of theoutput electromagnetic radiation is the intensity.

In a preferred embodiment, steps (C), (E) and (F) of the method of theinvention comprise:

-   -   (C) measuring a first interference pattern;    -   (E) measuring a second interference pattern; and    -   (F) relating the change from the first interference pattern to        the second interference pattern to the optical activity of the        stimulus of interest.

Preferably step (F) comprises: relating the intensity of at least a partof the first interference pattern relative to the intensity of at leasta part of the second interference pattern to the optical activity of thestimulus of interest. Particularly preferably the intensity is the totalintensity of substantially the whole of the interference pattern.

In a preferred embodiment, the method of the invention comprises:

-   -   (A1) providing an assembly as hereinbefore defined wherein the        means for relating the measurable response to the optical        activity of the stimulus of interest comprises:        -   an output polariser for orienting the degree of polarisation            of the output electromagnetic radiation;    -   (B) irradiating the optical component with incident        electromagnetic radiation of a first polarisation;    -   (C) measuring a first interference pattern;    -   (C1) orienting the output electromagnetic radiation into a third        degree of polarisation at which substantially the whole of the        first interference pattern has a minimum total intensity;    -   (D) introducing to the localised environment a stimulus of        interest;    -   (E) measuring a second interference pattern;    -   (E1) orienting the output electromagnetic radiation into a        fourth degree of polarisation at which substantially the whole        of the second interference pattern has a minimum total        intensity; and    -   (F1) relating the adjustment of the output polariser required to        orient the output electromagnetic radiation into the fourth        degree of polarisation to the optical activity of the stimulus        of interest.

In a preferred embodiment of the method, the input polariser is an inputplane polariser and the output polariser is an output plane polariser.

The method may further comprise the step of:

-   -   (G) deducing from the optical activity a structural,        conformational, biological or chemical property of the stimulus        of interest. For example, the conformational property may be the        chirality of the stimulus of interest or (where the stimulus of        interest is a protein) the folding of the protein.

Steps (A) to (F) of the method of the invention may be usefully repeatedat different wavelengths of incident electromagnetic radiation.

The present invention will now be described in a non-limitative sensewith reference to the accompanying Figures in which:

FIG. 1 illustrates schematically a first embodiment of the assembly ofthe invention;

FIG. 2 illustrates schematically a second embodiment of the assembly ofthe invention; and

FIG. 3 illustrates schematically a third embodiment of the assembly ofthe invention.

FIG. 1 illustrates schematically a first embodiment of the assembly ofthe invention. The assembly comprises a source of incidentelectromagnetic radiation 1, a lens arrangement 2, an input polariser(such as a prism) 3, a planar waveguide structure 4, an output polariser(an analyser) and prism 5, a lens arrangement 6 for focussing the outputelectromagnetic radiation and a photomultiplier tube 7 capable ofcounting photons 7. The polarisation of the incident electromagneticradiation relative to the planar waveguide structure 4 is determined bythe input polariser 3.

The planar waveguide structure 4 is of the type described inWO-A-98/22807 and in this embodiment comprises a silicon substrate layer4 a, silicon dioxide layers 4 b and 4 d, a silicon oxynitride layer 4 cand an absorbent layer 4 e. The silicon oxynitride layer 4 c acts as thereference waveguide and the absorbent layer 4 e acts as the sensingwaveguide.

The assembly is initialised by irradiating the planar waveguidestructure 4 with incident electromagnetic radiation of an appropriatewavelength from the source of incident electromagnetic radiation 1 inthe presence of materials which are not optically active and nulling theinput polariser 3 and output polariser 5. The incident electromagneticradiation is transmitted into the sensing waveguide 4e and the referencewaveguide 4 c simultaneously so that the level of radiation enteringeach is approximately the same. By ensuring that the materials in thelocalised environment 8 in contact with the sensing waveguide 4c are notoptically active, the output electromagnetic radiation will emerge fromthe planar waveguide structure 4 in the same orientation. The outputelectromagnetic radiation is coupled into free space thereby generatingan interference pattern at the photomultiplier tube 7. The outputpolariser 5 is aligned until the interference pattern of minimumintensity is detected by the photodetector 7. At this point, the inputpolariser 3 and output polariser 5 are mutually orthogonal.

The sensing waveguide 4 e is exposed in the localised environment 8 to asample (eg solution) containing the stimulus of interest which is eitheroptically active or has the capability to be optically active. Forexample, the sample may be brought into contact with or bound orimmobilised to the surface of the sensing waveguide 4 e. The planarwaveguide structure 4 is excited with the same wavelength ofelectromagnetic radiation as used during initialisation and the outputpolariser 5 is adjusted to regain the interference pattern of minimumintensity as determined by the photodetector 7. At this point theadjusted attitude of the output polariser 5 is directly proportional tothe optical activity of the stimulus of interest.

FIG. 2 illustrates schematically a second embodiment of the assembly ofthe invention. With the exception of the planar waveguide structure 4,the assembly is identical to the first embodiment described above withreference to FIG. 1. In this embodiment, the planar waveguide structurecomprises a silicon substrate layer 4 a, a silicon dioxide layer 4 b andan absorbent layer 4 c acting as the sensing waveguide. The assembly isoperated as described hereinbefore for the first embodiment.

FIG. 3 illustrates schematically a third embodiment of the assembly ofthe invention. The assembly comprises a source of electromagneticradiation 1, a lens arrangement 2, an input polariser (such as a prism)3, a polarisation maintaining fibre 5 with cladding 4, an outputpolariser (an analyser) 7 and prism, a lens arrangement 8 and aphotodetector 9. The cladding 4 has been partially removed to reveal alocalised environment 6 in which a stimulus of interest may be exposedto the surface of the polarisation maintaining fibre 5. The assembly isoperated as described hereinbefore for the first embodiment.

1. An assembly for measuring the optical activity of a stimulus ofinterest in a localised environment, said assembly comprising: a sourceof incident electromagnetic radiation; an input polariser for orientingthe incident electromagnetic radiation into a first degree ofpolarisation; an optical component capable of exhibiting a measurableresponse to a change in the degree of polarisation of the incidentelectromagnetic radiation into output electromagnetic radiation having asecond degree of polarisation caused by the introduction of or changesin the stimulus of interest in the localised environment; means formeasuring the measurable response; and means for relating the measurableresponse to the optical activity of the stimulus of interest.
 2. Anassembly as claimed in claim 1 wherein the input polariser is an inputplane polariser.
 3. An assembly as claimed in either of claims 1 or 2further comprising: a lens arrangement capable of focussing the incidentelectromagnetic radiation.
 4. An assembly as claimed in any of claims 1to 3 further comprising: a lens arrangement capable of focussing theoutput electromagnetic radiation.
 5. An assembly as claimed in anypreceding claim wherein the means for relating the measurable responseto the optical activity of the stimulus of interest comprises: an outputpolariser for orienting the degree of polarisation of the outputelectromagnetic radiation.
 6. An assembly as claimed in claim 5 whereinthe output polariser is an output plane polariser.
 7. An assembly asclaimed in any preceding claim wherein the optical component is apolarisation maintaining optical component.
 8. An assembly as claimed inany preceding claim wherein the optical component is an optical fibre.9. An assembly as claimed in any preceding claim wherein the measurableresponse is or relates to a change in the intensity of the outputelectromagnetic radiation.
 10. An assembly as claimed in any of claims 1to 8 wherein the optical component is a waveguide structure.
 11. Anassembly as claimed in claim 10 wherein the waveguide structure includeseither (1) one or more sensing layers capable of inducing in a secondarywaveguide a measurable response to a change in the degree ofpolarisation of the incident electromagnetic radiation into outputelectromagnetic radiation having a second degree of polarisation causedby the introduction of or changes in the stimulus of interest in thelocalised environment or (2) a sensing waveguide capable of exhibiting ameasurable response to a change in the degree of polarisation of theincident electromagnetic radiation into output electromagnetic radiationhaving a second degree of polarisation caused by the introduction of orchanges in the stimulus of interest in the localised environment.
 12. Anassembly as claimed in claim 10 or 11 wherein the waveguide structureincludes one or more sensing layers capable of inducing in a secondarywaveguide a measurable response to a change in the degree ofpolarisation of the incident electromagnetic radiation into outputelectromagnetic radiation having a second degree of polarisation causedby the introduction of or changes in the stimulus of interest in thelocalised environment.
 13. An assembly as claimed in claim 12 whereinthe optical component further comprises: an inactive secondary waveguidein which the sensing layer is substantially incapable of inducing ameasurable response to a change in the degree of polarisation of theincident electromagnetic radiation into output electromagnetic radiationhaving a second degree of polarisation caused by the introduction of orchanges in the stimulus of interest in the localised environment.
 14. Anassembly as claimed in claim 10 or 11 wherein the waveguide structureincludes a sensing waveguide capable of exhibiting a measurable responseto a change in the degree of polarisation of the incidentelectromagnetic radiation into output electromagnetic radiation having asecond degree of polarisation caused by the introduction of or changesin the stimulus of interest in the localised environment.
 15. Anassembly as claimed in claim 14 wherein the optical component furthercomprises: an inactive waveguide substantially incapable of exhibiting ameasurable response to a change in the degree of polarisation of theincident electromagnetic radiation into output electromagnetic radiationhaving a second degree of polarisation caused by the introduction of orchanges in the stimulus of interest in the localised environment.
 16. Anassembly as claimed in any of claims 10 to 15 wherein each waveguide ofthe waveguide structure is a planar waveguide.
 17. A method formeasuring the optical activity of a stimulus of interest in a localisedenvironment, said method comprising the steps of: (A) providing anassembly as defined in any preceding claim; (B) irradiating the opticalcomponent with incident electromagnetic radiation of a firstpolarisation; (C) measuring a first characteristic of the outputelectromagnetic radiation; (D) introducing to the localised environmenta stimulus of interest; (E) measuring a second characteristic of theoutput electromagnetic radiation; and (F) relating the change from thefirst characteristic of the output electromagnetic radiation to thesecond characteristic of the output electromagnetic radiation to theoptical activity of the stimulus of interest.
 18. A method as claimed inclaim 17 wherein the first and second characteristic of the outputelectromagnetic radiation is the intensity.
 19. A method as claimed ineither of claims 16 or 17 wherein steps (C), (E) and (F) comprise: (C)measuring a first interference pattern; (E) measuring a secondinterference pattern; and (F) relating the change from the firstinterference pattern to the second interference pattern to the opticalactivity of the stimulus of interest.
 20. A method as claimed in claim19 wherein step (F) comprises: relating the intensity of at least a partof the first interference pattern relative to the intensity of at leasta part of the second interference pattern to the optical activity of thestimulus of interest.
 21. A method as claimed in claim 20 wherein theintensity is the total intensity of substantially the whole of theinterference pattern.
 22. A method as claimed in any of claims 17 to 21comprising: (A1) providing an assembly as defined in any of claims 1 to16 wherein the means for relating the measurable response to the opticalactivity of the stimulus of interest comprises: an output polariser fororienting the degree of polarisation of the output electromagneticradiation; (B) irradiating the optical component with incidentelectromagnetic radiation of a first polarisation; (C) measuring a firstinterference pattern; (C1) orienting the output electromagneticradiation into a third degree of polarisation at which substantially thewhole of the first interference pattern has a minimum total intensity;(D) introducing to the localised environment a stimulus of interest; (E)measuring a second interference pattern; (E1) orienting the outputelectromagnetic radiation into a fourth degree of polarisation at whichsubstantially the whole of the second interference pattern has a minimumtotal intensity; and (F1) relating the adjustment of the outputpolariser required to orient the output electromagnetic radiation intothe fourth degree of polarisation to the optical activity of thestimulus of interest.
 23. A method as claimed in claim 22 wherein theinput polariser is an input plane polariser and the output polariser isan output plane polariser.
 24. A method as claimed in any of claims 17to 23 further comprising the step of: (G) deducing from the opticalactivity a structural, conformational, biological or chemical propertyof the stimulus of interest.
 25. A method as claimed in claim 24 whereinthe conformational property is the chirality of the stimulus of interestor (where the stimulus of interest is a protein) the folding of theprotein.